Bacillus cereus is a resilient foodborne pathogen, with biofilm formation and virulence regulated by quorum sensing (QS). This study evaluates GS-441524, the active metabolite of remdesivir, as an anti-virulence agent against B. cereus. At sub-inhibitory concentrations, GS-441524 dose-dependently reduced protease, lipase, hemolysin, and AI-2 production, impaired motility, and inhibited biofilm formation. Gene expression analysis showed downregulation of key QS (papR, luxS, plcR) and toxin (hblD, cytK, nprB) genes. Molecular docking indicated strong binding to HblD. In a Galleria mellonella model, GS-441524 enhanced survival upon B. cereus challenge. These findings demonstrate that GS-441524 disrupts QS and virulence, offering a promising strategy to control B. cereus.
Marine biofouling presents unique challenges in various industrial applications. This has historically been addressed using antifouling or fouling-release coatings. However, issues with the leaching of active or non-active components with non-target toxicity have resulted in increased regulatory scrutiny of these solutions. Natural polysaccharide-based coatings have emerged as an alternative as they contain many chemical functional groups that can enhance fouling-release/antifouling properties. In this review, polysaccharide structural properties, such as crystallinity and ionic character, are discussed with respect to functional coatings properties, such as wettability and antifouling performance. The limitations of polysaccharide antifouling performance in environments of high silt and salt are also noted. Mitigation of these challenges by hydrophobic chemical modifications - amphiphilic polysaccharides - are explored as the frontier for high-performance coatings development. Recommendations are provided for future coatings development to better reflect real-world conditions, including using polysaccharides in commercial resin technologies and more extensive field test validation.
The current study describes the evaluation of 16 synthetic linear tripeptides combining structural elements from the innate defence system with brominated amino acids often found in antifouling compounds isolated from sessile marine organisms. These organisms have clean exteriors and their chemical arsenal of natural antifoulants is interesting from an antifoulant design perspective. The peptides were evaluated for their ability to inhibit the settlement and metamorphosis of the soft and hard macrofoulers Ciona savignyi and Amphibalanus improvisus which are two key macrofoulers challenging to remove once established. It was shown that the evaluated compounds were generally highly active antifoulants with low micromolar IC50-values against both fouling organisms via different mechanisms. Several of the compounds were shown to be biocidal against Ciona savignyi larvae at the lowest evaluated concentration (1 µM). Against Amphibalanus improvisus, a non-biocidal antifouling effect was generally observed with the most potent compound displaying an IC50-value of 0.5 µM. The study illustrated how these two groups of natural compounds combined are highly bioactive and can yield highly efficient antifoulants.
Gloeocapsa spp. are a type of airborne cyanobacteria that are known for causing black streaks on asphalt roofs. Embedding copper containing granules into asphalt shingles was developed to prevent cyanobacterial growth on roof shingles. There are currently no established methods for testing the anti-cyanobacterial activity of copper roofing granules. First, the minimum inhibitory concentration (MIC) of Cu2+ ions in solution was evaluated by monitoring growth in a 96 well plate over 14 days. Using a Gompertz function the MIC of Cu2+ was determined to be 0.592 ppm for G. alpicola. To test copper granules used in shingles, the granules were either autoclaved in BG11 media or sterilized using anhydrous ethanol, dried, and added to sterile BG11. Once inoculated with G. alpicola, growth was monitored using optical density (OD) and fluorescence measurements. Debris from the granules interfered with OD measurements but fluorescence clearly showed granules formulated with copper prevented cyanobacterial growth and those without did not. This work is of practical use for companies looking to formulate anti-cyanobacterial products like roof granules as it reports a specific MIC for this airborne species of cyanobacteria often found on roofs and also validates a methodology for testing roof granules.
Within seconds of placement, an adsorbed conditioning film replaces the implant's manufactured substrate and becomes the true interface host cells and microorganisms encounter. Conventional strategies engineered to kill or suppress bacteria therefore act on an interface neither party meets, helping explain the inconsistent clinical performance of broadly bactericidal designs. We propose interfacial selectivity, the capacity of a surface to favor beneficial over detrimental interactions, as a unifying design framework for implant biomaterials, and show how interfacial physicochemistry can be programmed to discriminate between colonizers. The implant boundary is viewed as three coupled interfaces: implant-film, film-microbiome, and film-host. Shared drivers, surface energy, charge, hydration, and multiscale topography, govern all three, so modifications intended to deter pathogens also influence commensal colonization, soft-tissue sealing, and osteoimmune balance. A regime map of interfacial forces reinterprets functionalization strategies within a common selectivity space, and a proposed selectivity index, applied here to published data for three archetypal surfaces, reorients evaluation from short-term killing toward the balance between host-beneficial and pathogen-beneficial outcomes. We derive transferable design principles, propose a minimum reporting standard, identify methodological gaps limiting translation, and argue that genomic and multi-omic calibration could enable patient-specific implementation across dental, percutaneous, and orthopedic implants.
Acinetobacter baumannii is a formidable nosocomial pathogen whose biofilm production is a key contributor to its multidrug resistance. This study reveals the significant antibacterial and antibiofilm potential of Salvia abrotanoides essential oils (EOs) against MDR and XDR A. baumannii. GC-MS analysis identified 19 novel compounds within the leaf (L-EO) and flower (F-EO) extracts. F-EO exhibited greater potency, inhibiting bacterial growth and biofilm formation at 1.75 mg/mL and 0.9 mg/mL, respectively. At sub-inhibitory concentrations, both EOs caused substantial downregulation (>84%) of the critical biofilm genes bap and csuD, which was visually confirmed by SEM. Cytotoxicity assays on A549 cells showed IC50 values of 1.6 mg/mL (L-EO) and 2.3 mg/mL (F-EO). These findings position S. abrotanoides EOs as a promising therapeutic strategy to combat resilient, biofilm-mediated infections.
Microplastics in coastal waters provide persistent surfaces for microbial colonization and biofilm formation, supporting complex microbial assemblages known as the plastisphere. This study examines the plastisphere-biofouling nexus through a PRISMA-guided systematic review, bibliometric mapping, narrative-quantitative synthesis, and conceptual evidence integration of studies published between 2015 and 2025. From 1,248 records identified in Web of Science Core Collection and Scopus, 124 primary studies were retained for comparative synthesis. The reviewed evidence shows that microplastics frequently support distinct biofilm-associated microbial communities, although reported patterns vary across polymer type, exposure duration, environmental setting, microbial method, and biofilm measurement approach. Proteobacteria and Bacteroidetes were repeatedly reported among dominant bacterial groups, while opportunistic genera such as Vibrio and Pseudoalteromonas were detected in some plastisphere biofilms. Evidence for polymer-specific microbial diversity, contaminant retention, and environmental-driver effects remains context-dependent. Biofilm-coated microplastics may modify contaminant interactions under specific conditions, but current evidence does not support universal contaminant enhancement, confirmed pathogen transmission, or direct human health risk. This review identifies the plastisphere-biofouling nexus as an emerging ecological interface in coastal waters and highlights the need for standardized, field-relevant, and functionally validated studies.
Multidrug-resistant (MDR) Pseudomonas aeruginosa infections are difficult to treat due to biofilm formation. This study evaluates the antimicrobial and antibiofilm efficacy of murepavadin (MUR), a novel peptidomimetic targeting the LptD protein, alone and combined with conventional antibiotics against clinical isolates. The minimum inhibitory concentrations (MICs) of 50 isolates against MUR and five antibiotics were determined. Synergy in planktonic cultures was assessed by checkerboard assays. For biofilm, the Bliss independence model was applied using MBIC (minimum biofilm inhibitory concentration) and the concentration yielding ≥90% biomass reduction, as measured by crystal violet (CV) staining. MUR showed potent activity against planktonic cells (MIC50/90: 0.25/0.5 mg l-1). Synergy tests revealed consistent MUR-colistin synergy across all strains, with notable interactions against carbapenemase-producers when combined with meropenem. In biofilms, MUR demonstrated significantly lower MBIC and lower concentrations required for ≥90% biomass reduction (CV staining). Bliss analysis confirmed strong synergistic biofilm inhibition in all combinations, most pronounced with tobramycin. MUR exhibits high efficacy against both planktonic and biofilm-forming MDR P. aeruginosa. Its synergy with colistin and tobramycin highlights its potential as a strategic combination partner.
Natural deep eutectic solvents (NADES) have recently emerged as promising 'green' antibiofilm agents due to their ability to solubilize biological macromolecules. In this study, three chemically distinct NADES formulations were evaluated against Pseudomonas fluorescens WCS365 and Staphylococcus epidermidis ATCC 35984. The tested formulations choline chloride-lactic acid (CCLA), choline chloride-urea (CCU), and choline chloride-xylitol (CCX) were assessed for their effects on planktonic growth, biofilm formation, and mature biofilms. All NADES showed moderate inhibition of planktonic growth, while biofilm formation was significantly reduced in a formulation- and species-dependent manner, with CCLA displaying the strongest activity. Treatment of pre-formed 24 h biofilms resulted in partial but significant biomass reduction, reaching up to ∼69% for P. fluorescens and ∼51% for S. epidermidis. Confocal microscopy confirmed pronounced structural disruption of mature biofilms following NADES exposure, particularly for CCLA. These findings highlight the potential of organic acid-based NADES as biocides, which are more environmentally compatible than conventional biocides.
Candida albicans (C. albicans) biofilms exhibit markedly enhanced tolerance to conventional antifungal agents, necessitating the identification of novel therapeutic alternatives. Chrysin, a naturally occurring flavonoid distributed in propolis and Passiflora species, possesses well-documented antioxidant and anti-inflammatory properties. However, its activity against C. albicans biofilms remains uncharacterized. This study comprehensively evaluated the antibiofilm efficacy of chrysin in vitro and in vivo. The sessile minimum inhibitory concentration causing 50% inhibition (SMIC50) of chrysin against C. albicans SC5314 was 128 μg/mL. Chrysin suppressed biofilm metabolic activity, reduced biomass, and decreased cell viability in a concentration-dependent manner, as demonstrated by XTT reduction assay, crystal violet staining, and live/dead fluorescence staining. Scanning electron microscopy (SEM) revealed progressive dismantling of hyphal networks and disruption of biofilm architecture. Chrysin significantly elevated intracellular reactive oxygen species (ROS) levels and induced mitochondrial membrane potential (MMP) depolarization in a dose-dependent manner, indicating that oxidative stress induction is a key antifungal mechanism. In a murine oral candidiasis model, chrysin reduced fungal burden and tongue lesion scores in a concentration-dependent manner, with the 256 μg/mL group achieving outcomes comparable to fluconazole. Chrysin also attenuated pro-inflammatory cytokines interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α), while increasing interleukin-10 (IL-10). Biosafety assessment demonstrated cell viability above 80% in human oral keratinocytes (HOK) and above 85% in mouse fibroblasts (L929) at concentrations up to 512 μg/mL, with no observable organ toxicity in vivo. Collectively, these findings establish chrysin as a potent, mechanistically defined, and safe natural antifungal candidate with promising translational potential for the management of C. albicans biofilm-associated infections.
Recent studies have shown ultraviolet light (UVC) to provide increased biofouling performance when combined with a marine coating, however, it is unknown what impacts this may have on the coating. In this two-part study, we address the impact of UVC exposure on a silicone fouling release coating, as a function of frequency and dosage (4.97 J/cm2 to 7.15 kJ/cm2). Higher doses were found to alter the coating properties (e.g. color, hardness, surface energy, adhesion properties). They also resulted in a greater relative abundance of aliphatic (C-C) and siloxane (Si-O-Si) and a decrease in carboxyl (C-O) moieties, along with an increase in the reduced modulus of the coating. However, when paired with a marine coating, UVC will not need to be applied in these high doses. The changes observed with lower doses used were minimal, suggesting that the combination of UVC and marine coatings may prolong the efficacy of biofouling prevention.
This study examines the antibacterial and antibiofilm activities of essential oils from Thymus vulgaris, Rosmarinus officinalis, and Lavandula angustifolia. Chemical analysis identified carvacrol, linalool, and camphor as key constituents contributing to activity against E. coli, S. aureus, and P. aeruginosa. T. vulgaris oil, rich in phenolic and flavonoid content, showed the strongest efficacy. R. officinalis and L. angustifolia oils also demonstrated notable antimicrobial effects, reducing bacterial viability, biofilm biomass, and metabolic activity, although effectiveness varied with species and biofilm structure. T. vulgaris oil effectively removed E. coli and P. aeruginosa biofilms at 2 × MIC (log reduction 7) but was less effective against S. aureus (log reduction 1.7) at 2 × MIC. The activity of these oils may be associated with disruption of bacterial membranes, biofilm architecture, and induction of oxidative stress. Mammalian toxicity at 3 × MIC shows a low toxic profile for T. vulgaris and R. officinalis, but a high toxic profile for L. angustifolia. These findings highlight T. vulgaris and R. officinalis essential oils as promising antibacterial agents in terms of both efficacy and safety.
Berberine, a naturally occurring isoquinoline alkaloid, has demonstrated strong antibiofilm properties against Staphylococcus aureus. However, the molecular mechanisms underlying this activity remain poorly understood. Here, we report that berberine effectively inhibits S. aureus biofilm formation with a minimum biofilm inhibitory concentration (MBIC) of 32 µg ml-1. XTT assays demonstrated a marked reduction in biofilm metabolic activity, while high-content screening (HCS) combined with three-dimensional reconstruction revealed significant decreases in biofilm biomass and thickness. Mechanistically, berberine selectively suppressed the production of key extracellular polymeric substance (EPS) components, particularly polysaccharide intercellular adhesin (PIA) and proteins, without significantly affecting extracellular DNA (eDNA) levels. Transcriptomic profiling integrated with protein-protein interaction (PPI) network modeling suggested that srrA, a response regulator within the two-component system, may serve as a central hub gene downregulated by berberine. Quantitative PCR (qPCR) further corroborated the altered expression of srrA and associated genes (walR, atpB, luxS, icaR). Collectively, these findings provide mechanistic insights into berberine's antibiofilm activity and point to srrA-mediated EPS regulation as a candidate pathway warranting further investigation.
Candidozyma auris (formerly Candida auris) (C. auris), an emerging multidrug-resistant fungal pathogen, forms biofilms as a virulence factor. This study aimed to determine the effect of phenotypic switch on C. auris biofilm formation and virulence gene expression in mono- and co-culture with Staphylococcus aureus. Phenotypic switching was induced by prolonged incubation, and biofilms were developed in RPMI-1640, YEPD, SDB, and BHIYE. The biofilm biomass and total cell count were measured. SAP5 and ALS5 gene expression was quantified using qPCR. The 4th switched generation mono-culture biofilm in BHIYE produced the highest biomass (3.34 ± 0.08) and total cell count (5.66 ± 0.03 log10 cells mL-1). In addition, SAP5 and ALS5 expression peaked in the 2nd switched generation mono-culture by 10.43 ± 0.44-fold and 4.764 ± 0.01-fold, respectively. Co-culture biofilms exhibited significantly higher ALS5 expression in selected switched generations compared to unswitched C. auris (p < 0.05). In conclusion, phenotypic switching enhanced biofilm formation and modulated the expression of SAP5 and ALS5 in C. auris.
In orthodontic treatment, insufficiently cleaned clear-aligners can serve as reservoirs for biofilms. This study evaluates in-vitro biofilm models and investigates the antibiofilm efficacy of boron-based formulations with natural substances. Minimum inhibitory concentrations (MICs) were determined by microdilution, and Fractional inhibitory concentration by checkerboard methods. Biofilm inhibition was assessed at sub-MIC levels, and mature biofilm-eradication at supra-MICs. Time-kill assays were performed on mixed-biofilms of Streptococcus mutans and Candida albicans on clear-aligners. The MIC values of boric acid, sodium borate, and natural substances against S. mutans were 1,250-5,000, 4,000-8,000, and 2.5->10,000 μg/mL, respectively. Synergistic combinations included eucalyptol, epigallocatechin-gallate, resveratrol, and cinnamaldehyde. Biofilm inhibition rates ranged from 49%-58%, and eradication concentrations were 8-16xMIC. A formulation containing boric acid reduced viable cells by ≥5-log10 in time-kill assays. These findings indicate that boron-based formulations with natural substances suggest notable antibiofilm activity against mixed-species biofilms and may provide preliminary in vitro data for aligner-cleaning or oral-care products.
Under resource-limited conditions, biofilms display highly complex competitive behaviors and distinct growth dynamics. This study investigated Bacillus subtilis strain engineered with three fluorescent reporter genes by integrating microscopic and macroscopic analyses to uncover the intrinsic mechanisms driving heterogeneous growth under competitive constraints. The results show that stress accumulation leads to the formation of a pronounced thickness peak in the interfacial region between two neighboring biofilms under strong competition (inoculation-center distance, d = 2 mm). Under nutrient-rich conditions, a ring-shaped elevation emerges approximately 4 mm from the colony center, providing an expanded spatial niche that facilitates bacterial survival. Self-healing rates of incisions at different locations of biofilms were compared to evaluate the effect of competition. The findings show that the self-healing capacity of biofilms in competitive environments is governed by both biofilm age and competitive intensity. Incorporating nutrient diffusion, environmental pressure, and phenotype-switching probabilities, a cellular automaton model was further developed to simulate the competitive growth of the biofilms.
Staphylococcus aureus biofilms pose significant challenges in medical and industrial settings by exhibiting high resistance to conventional treatments. Although cold atmospheric plasma (CAP) shows promise as an antibacterial agent, the mechanisms of interaction with bacteria, particularly in saline environments, remain poorly understood. S. aureus biofilms were exposed to Argon CAP, and morphological and metabolic changes were evaluated using confocal laser scanning microscopy, field emission scanning electron microscopy, Fourier transform infrared spectroscopy, atomic force microscopy, and XTT fluorescence. Planktonic cells were further examined by transmission electron microscopy. The findings revealed significant biofilm disruption with 94% metabolic inactivation after 300 s. To distinguish direct plasma effects, plasma-treated saline was tested and demonstrated to inactivate within 30 min. Optical emission spectroscopy detected intense Na emission (589 nm) during plasma-saline interactions; however, this observation is reported as a spectral feature rather than a mechanistic conclusion, and further experiments are required to clarify its potential relevance.
Acid-producing bacteria (APB) play significant role in pipeline corrosion by producing corrosive metabolites. Current APB mitigation strategies are often unsuccessful, costly, or not environmentally friendly. In this study, nitrate was used to manipulate microbial interaction and control APB activities. Microbial diversity was manipulated by subsampling and culturing sufficient replicates of serially diluted subsamples to achieve random distribution in the absence or presence of nitrate. Microbial activities were evaluated by monitoring changes in pH, corrosion rate, production of organic acids, changes in community composition, and functional gene abundance. The study identified a change in microbial community assemblage with decreased abundance of Clostridium upon nitrate addition, resulting in limited APB activities such as acid production and biocorrosion. Nitrate resulted in about a 3.5-fold decrease in carbon steel weight loss corrosion induced by APB but did not affect microbial growth and abundance of metabolic genes, suggesting that the inhibitory effect is likely due to changes in redox potential of culture media or increased growth of APB competitors. The study also identified the importance of nitrate as an economically viable and environmentally sustainable approach to mitigate APB-induced biocorrosion.
Barnacles are a highly diverse group that includes sessile, stalked, and parasitic forms, all capable of adhering strongly to wet surfaces. While acorn barnacle adhesion has been well studied, the mechanisms in stalked barnacles remain poorly understood. Herein, the adhesive of Capitulum mitella is investigated, revealing a two-phase secretion: a fluid-phase mixture extruded through glandular structures and a solidified adhesive with distinct lamellar architecture. The outer layers of adhesive can resist environmental stress, while the inner fibrous network contributes to cohesion and viscoelasticity. These components differ markedly in morphology and chemical composition. Raman spectroscopy, FTIR, and thioflavin T staining confirm the absence of amyloid-like β-sheet structures, in contrast to previously reported amyloid-based architectures in acorn barnacle adhesives. The results highlight species-specific adhesive strategies in stalked barnacles and offer insights relevant to the development of bioinspired adhesives and antifouling technologies.
The indiscriminate use of antibiotics and biofilm formation have contributed to the emergence of resistant infections. Natural compounds such as limonene and its derivatives have gained attention due to their therapeutic potential and lower risk of resistance. This study evaluated the antibacterial and antibiofilm activity of limonene isomers ((R)- and (S)-limonene) and their derivatives ((S)-perillyl alcohol and (S)-perillaldehyde), as well as their combined effects with β-lactam antibiotics against Gram-positive and Gram-negative strains of clinical and food relevance. Disk diffusion, MIC, MBC, growth curve, biofilm quantification (biomass and cell viability), and checkerboard assays were performed. (S)-limonene, (S)-perillyl alcohol, and (S)-perillaldehyde showed bactericidal activity, whereas (R)-limonene was bacteriostatic. Structural and electronic analyses indicated that oxygenation and substituents modify electron density, polarity, and reactivity, influencing biological activity. All compounds delayed bacterial growth and significantly reduced biofilm biomass and cell viability, with S-isomers achieving reductions of up to 90%. Combinatory analysis revealed synergistic and additive effects with oxacillin, benzylpenicillin, ampicillin, and ceftriaxone, particularly against MRSA and Klebsiella pneumoniae. In silico docking suggested potential inhibition of Gyrase B, thymidylate kinase, and topoisomerase through interactions with active-site residues. ADMET predictions indicated favorable permeability and absorption, especially for S-PA and R-LMN. These findings highlight the potential of limonene derivatives as alternative or adjunct antimicrobial agents against biofilm-associated infections.
